Conventional nanofiltration or reverse osmosis water filtration membranes have been known for many decades. Typically, they are made by casting a support membrane (often polysulfone or polyethersulfone); immersing the resulting cast in an aqueous solution of a diamine; removing excess from the surface of the membrane; immersing the membrane in an organic solution of a trifunctional acyl halide; and curing the resulting product to produce a polyamide layer. Washing and secondary coating are then carried out as necessary.
It is known from WO 01/32146 that membrane proteins may be incorporated into the walls of vesicles made from amphiphilic ABA block copolymers. This document includes extensive discussion of the nature of the polymers, and discloses that the polymers may have polymerizable groups at both chain ends. These polymerizable groups can be polymerized after the formation of the self-assembled vesicles, the polymerisation occurring exclusively intravesicularly. WO 2004/011600 discloses that aquaporins may be incorporated into triblock co-polymers to form a membrane which will only pass water, excluding all contaminants. Since these disclosures, much work has been carried out to develop commercially viable membranes incorporating transmembrane proteins, and particularly water filtration membranes based on aquaporins. The challenge is to produce a working membrane, which is physically sufficiently robust to withstand the necessary conditions. WO 2009/076174 describes a method of preparing substantially flat membranes based on block copolymers and aquaporins. According to Zhao et al, J. Membrane Sci. 2012, 422-428, various proposed methods of producing aquaporin membranes include polymer tethered bio-layers, biomembrane aperture partition arrays, membrane supported lipid bilayer via vesicle fusion, and vesicles suspended over membrane pores, but most of these are not able to withstand the high hydrostatic pressure that is required. Zhao's own solution to the problem is in effect to use a conventional membrane preparation as described above, modified by addition of aquaporin-loaded lipid vesicles (i.e. liposomes) to the aqueous solution of diamine. The result provides liposomes embedded in a polyamide layer. Although Zhao reports the results obtained positively, it is clear from the data provided that although a small increase of water flux is obtained (FIG. 4(a)) no enhancement of the ability of the membrane to reject solute is found compared with conventional membranes (FIG. 5). It is believed that this is because the aquaporin-loaded liposomes become completely surrounded by polyamide, and thus the primary water flux through the membrane is via the polyamide (i.e. via the conventional path of the base membrane), and only partially through the aquaporin channels. WO 2013/043118, also from Zhao et al, describes the same technology and also discloses that block copolymers can be used to form vesicles, either containing or not containing aquaporins, and embedded in a polyamide layer. Again, the results plainly show that water flux via the polyamide layer and not exclusively via the aquaporin channels is obtained.
Xie et al, J. Mater. Chem A, 2013, 1, 7592 describes a process comprising (i) incorporating aquaporin into self-assembled polymer vesicles based on a polymer primarily (95%) having methacrylate end groups but also containing some (3%) carboxylic acid end groups; (ii) cross-linking the methacrylate end groups using UV light; (iii) depositing and covalently immobilizing the cross-linked vesicles on a support in such a concentration that isolated vesicles are disposed separately from each other on the surface of the support; and (iv) creating a thin polymer layer between the individual vesicles by the process known as “surface imprinting”. In this process, it is important that the size of the immobilized vesicles is such that they are larger than the thickness of the imprinted polymer layer to prevent blockage of the aquaporin water channels. The process is said to exhibit high mechanical strength and stability during water filtration, but it is also stated that the most critical issue is that the imprinted polymer layer was not sufficiently dense to prevent all of the solute and water molecules from permeating. Further, only very limited flow rates are obtainable by such a system.
Accordingly, there still remains a need for a process which leads to a physically robust membrane incorporating transmembrane proteins, particularly a membrane which uses aquaporins acting effectively for water filtration. Our copending application ref. no. P021889WO claiming priority from GB 1405390 relates to such a membrane: that invention provides a filtration membrane which comprises a porous support and, covalently bonded to a surface thereof, a layer comprising a plurality of vesicles having transmembrane proteins incorporated therein, said vesicles being formed from an amphiphilic block copolymer, characterised in that within said layer, vesicles are covalently linked together to form a coherent mass.
The propensity of known amphiphilic polymers to form vesicles, rather than other self-assembly structures such as miscelles, is known to depend on the absolute and relative sizes of the hydrophobic and hydrophilic blocks. Previously, the nature of these blocks has been believed to be much the most important factor determining the ease with which vesicles can be formed. Polymers with a number of different end groups have been used in vesicle formation, but no effect on vesicle formation has been noted. For example, US 2008/0305149 discloses PMOXA-PDMS-PMOXA block copolymers having —OH, —NH2, —NH-piperazine, —SH and —COONa end groups. Surprisingly, we have now found that the presence of an end group including both —NH2 and —NH— groups, i.e. which includes both primary and secondary amine groups, makes a major difference, and the use of (poly)2-C1-3alkyl-2-oxazoline, especially (poly)2-methyl-2-oxazoline/(poly)dimethyl siloxane block copolymers having at least one such end group has proved particularly valuable for the preparation of vesicles.